Surfactant protein A modulates the inflammatory response in macrophages during tuberculosis

Abstract

Tuberculosis leads to immune activation and increased human immunodeficiency virus type 1 (HIV-1) replication in the lung. However, in vitro models of mycobacterial infection of human macrophages do not fully reproduce these in vivo observations, suggesting that there are additional host factors. Surfactant protein A (SP-A) is an important mediator of innate immunity in the lung. SP-A levels were assayed in the human lung by using bronchoalveolar lavage (BAL). There was a threefold reduction in SP-A levels during tuberculosis only in the radiographically involved lung segments, and the levels returned to normal after 1 month of treatment. The SP-A levels were inversely correlated with the percentage of neutrophils in BAL fluid, suggesting that low SP-A levels were associated with increased inflammation in the lung. Differentiated THP-1 macrophages were used to test the effect of decreasing SP-A levels on immune function. In the absence of infection with Mycobacterium tuberculosis, SP-A at doses ranging from 5 to 0.01 micro g/ml inhibited both interleukin-6 (IL-6) production and HIV-1 long terminal repeat (LTR) activity. In macrophages infected with M. tuberculosis, SP-A augmented both IL-6 production and HIV-1 LTR activity. To better understand the effect of SP-A, we measured expression of CAAT/enhancer binding protein beta (C/EBPbeta), a transcription factor central to the regulation of IL-6 and the HIV-1 LTR. In macrophages infected with M. tuberculosis, SP-A reduced expression of a dominant negative isoform of C/EBPbeta. These data suggest that SP-A has pleiotropic effects even at the low concentrations found in tuberculosis patients. This protein augments inflammation in the presence of infection and inhibits inflammation in uninfected macrophages, protecting uninvolved lung segments from the deleterious effects of inflammation.

FIG. 1.

SP-A levels were reduced in TB patients and returned to normal after 1 month of antituberculous therapy. BALF from normal individuals and from radiographically involved and uninvolved lung segments of subjects with TB were assayed for SP-A by a commercially available ELISA. (A) SP-A levels were reduced in the involved lung segments of TB subjects compared to the levels in uninvolved lung segments and normal individuals. (B) SP-A levels increased after 1 month of antituberculous therapy. (C) Total phosphorus levels from BALF of all subjects. Data are the means; error bars are SEM.

FIG. 2.

SP-A levels were inversely correlated with the BALF neutrophil count. (A) SP-A levels were inversely correlated with the PMN content in BALF from normal individuals and involved and uninvolved lung segments. (B) SP-A levels were inversely correlated with the involved lung segments of TB subjects.

FIG. 3.

Low levels of SP-A increased proinflammatory cytokine production in vitro. THP-1 macrophages were infected with either BCG or H37Ra with or without SP-A for 24 h. (A) IL-6 production was increased in infected THP-1 macrophages (M.TB+) at doses as low as 0.01 μg/ml, and the opposite effect was observed in uninfected cells (M.TB−) (n = 5) (B) HIV LTR activity was assayed with a CAT reporter construct. HIV LTR activity was increased in infected THP-1 macrophages at doses as low as 0.01 μg/ml, and the opposite effect was observed in uninfected cells. The asterisk indicates that the P value is 0.02 for a comparison with SP-A-treated uninfected cells.

FIG. 4.

Low levels of SP-A increase the C/EBPβS/I ratio. THP-1 macrophages were infected with H37Ra or BCG with or without SP-A for 24 h. (A)Representative immunoblot for C/EBPβ. Infection with mycobacteria increased the levels of both isoforms (compare lanes 1 and 3). SP-A decreased the amount of the inhibitory isoform, and themaximal effect was observed at 0.01 μg/ml (compare lanes 3 and 7). In contrast, a similar dose of SP-A had no effect on C/EBPβ levels in uninfected cells (lanes 1 and 2). M.Tb, M. tuberculosis. (B) Immunoblots from three or four separate experiments were quantified by densitometry. The results are expressed as the fold changesin the S/I ratio relative to non-SP-A-treated controls. SP-A at a dose of 0.01 μg/ml increased the C/EBPβS/I ratio, while SP-A had little effect in uninfected cells. All lanes were normalized for total protein content (50 μg).−M.TB, without M. tuberculosis; +M.TB, with M. tuberculosis.